The mechanisms underlying caffeine’s acute effects and withdrawal symptoms are not entirely understood. The purpose was to determine whether the clusters of acute effects or withdrawal symptoms are associated with genetic polymorphisms in DARPP-32 and COMT, which mediate some of caffeine’s physiological effects. Subjects (n=1135) were from the Toronto Nutrigenomics and Healthy Study. Fourteen well-described acute effects of caffeine co-exist in six groups, while fourteen well-characterized withdrawal symptoms co-exist in three groups. Neither the rs907094 C>T polymorphism in the PPP1R1B gene encoding DARPP-32, nor the COMT Val158Met affected the odds of reporting any acute effects or withdrawal symptoms cluster. Among individuals consuming ≥ 200 mg/d of caffeine, Met/Met homozygotes were more likely to report the “increased heart rate” acute effects cluster. These results suggest that ‘slow’ COMT activity, conferred by the Met allele, may explain part of the inter-individual variability in the risk for increased heart rate among heavy caffeine consumers.

The mechanisms underlying caffeine’s acute effects and withdrawal symptoms are not entirely understood. The purpose was to determine whether the clusters of acute effects or withdrawal symptoms are associated with genetic polymorphisms in DARPP-32 and COMT, which mediate some of caffeine’s physiological effects. Subjects (n=1135) were from the Toronto Nutrigenomics and Healthy Study. Fourteen well-described acute effects of caffeine co-exist in six groups, while fourteen well-characterized withdrawal symptoms co-exist in three groups. Neither the rs907094 C>T polymorphism in the PPP1R1B gene encoding DARPP-32, nor the COMT Val158Met affected the odds of reporting any acute effects or withdrawal symptoms cluster. Among individuals consuming ≥ 200 mg/d of caffeine, Met/Met homozygotes were more likely to report the “increased heart rate” acute effects cluster. These results suggest that ‘slow’ COMT activity, conferred by the Met allele, may explain part of the inter-individual variability in the risk for increased heart rate among heavy caffeine consumers.

In silico studies of the effect of phenolic compounds from grape seed extracts on the activity of phosphoinositide 3-kinase (PI3K) and farnesoid X receptor (FXR)Montserrat Vaqué Marquès En aquesta tesis es pretén aplicar metodologies computacionals (generació de farmacòfors i docking proteïna lligand) en l'àmbit de la nutigenòmica (ciència que pretén entendre, a nivell molecular, com els nutrients afecten la salut). S'aplicaran metodologies in silico per entendre a nivell molecular com productes naturals com els compostos fenòlics presents en la nostra dieta, poden modular la funció d'una diana comportant un efect en la salut. Aquest efecte es creu que podria ser degut a la seva interacció directa amb proteïnes de vies de senyalització molecular o bé a la modificació indirecta de l'expressió gènica. Donat que el coneixement de l'estructura del complex lligand-receptor és bàsic per entendre el mecanisme d'acció d'aquests lligands s'aplica la metodologia docking per predir l'estructura tridimensional del complex. En aquest sentit, un dels programes de docking és AutoGrid/AutoDock (un dels més citats). No obstant, l'automatització d'AutoGrid/AutoDock no és trivial tan per (a) la cerca virtual en una llibreria de lligands contra un grup de possibles receptors, (b) l'ús de flexibilitat, i (c) realitzar un docking a cegues utilitzant tota la superfície del receptor. Per aquest motiu, es dissenya una interfície gràfica de fàcil ús per utilitzar AutoGrid/AutoDock. Blind Docking Tester (BDT) és una aplicació gràfica que s'executa sobre quatre programes escrits en Fortran i que controla les condicions de les execucions d'AutoGrid i AutoDock. BDT pot ser utilitzat per equips d'investigadors en el camp de la química i de ciències de la vida interessats en dur a terme aquest tipus d'experiments però que no tenen suficient habilitats en programació. En la modulació del metabolisme de la glucosa, treballs in vivio i in vitro en el nostre grup de recerca s'han atribuït els efectes beneficiosos de l'extracte de pinyol de raïm en induir captació de glucosa (punt crític pel manteniment de l'homeostasis de la glucosa). No obstant alguns compostos fenòlics no tenen efecte en la captació de la glucosa, d'altres l'inhibeixen reversiblement. En alguns casos aquesta inhibició és el resultat de la competició dels compostos fenòlics amb ATP pel lloc d'unió de l'ATP de la subunitat catalítica de la fosfatidil inositol 3-kinasa (PI3K). Estudis recents amb inhibidors específics d'isoforma han identificat la p110α (la subunitat catalítica de PI3Kα) com la isoforma crucial per la captació de glucosa estimulada per insulina en algunes línies cel·lulars. Els programes computacionals han estat aplicats per tal de correlacionar l'activitat biològica dels compostos fenòlics amb informació estructural per obtenir una relació quantitativa estructura-activitat (3D-QSAR) i obtenir informació dels requeriments estructura-lligand per augmentar l'afinitat i/o selectivitat amb la diana (proteïna). Tot hi haver-se demostrat que l'adició d'extractes de compostos fenòlics en l'aliment pot tenir en general un benefici per la salut, s'ha de tenir en compte que l'estudi 3D-QSAR (construït a partir d'inhibidors sintètics de p110α) prediu que algunes d'aquestes molècules poden agreujar la resistència a la insulina en individus susceptibles dificultant la capatació de glucosa en múscul i teixit adipós i, per tant, produir un efecte secundari indesitjat. Resultats en el nostre grup de recerca han demostrat que compostos fenòlics presents en extractes de llavor de raïm incrementen l'activitat del receptor "farnesoid x receptor" (FXR) de manera dosi depenent quan el lligand natural de FXR (CDCA) és present. Les metodologies in silico, docking i 3D-QSAR, han estat aplicades juntament amb dades biològiques d'agonistes no esteroidals de FXR que s'uneixen a un lloc d'unió proper però diferent al lligand esteroidal 6CDCA. Els resultats han mostrat que els compostos fenòlics no són capaços d'activar FXR per ells mateixos però poden afegir noves interaccions que estabilitzarien la conformació activa de FXR en presència del lligand natural CDCA. Els compostos fenòlics podrien induir canvis conformacionals específics que augmentarien l'activitat de FXR. In silico studies of the effect of phenolic compounds from grape seed extracts on the activity of phosphoinositide 3-kinase (PI3K) and farnesoid X receptor (FXR)Montserrat Vaqué Marquès This thesis was written with the aim of applying computational methods that have already been developed for molecular design and simulation (i.e. pharmacophore generation and protein-ligand docking) to nutrigenomics. So, in silico tools that are routinely used by the pharmaceutical industry to develop drugs have been used to understand, at the molecular level, how natural products such as phenolic compounds (i.e. molecules that are commonly found in fruits and vegetables) can improve health and prevent diseases. Therefore, we first focused on predicting the structure of protein-ligand complexes. The docking algorithms can use the individual structures from receptor and ligand to predict (1) whether they can form a complex and (2) if so, the structure of the resulting complex. This prediction can be made, for instance, with AutoGrid/AutoDock, the most cited docking software in the literature. The automation of AutoGrid/AutoDock is not trivial for tasks such as (1) the virtual screening of a library of ligands against a set of possible receptors; (2) the use of receptor flexibility and (3) making a blind-docking experiment with the whole receptor surface. Therefore, in order to circumvent these limitations, we have designed BDT (i.e. blind-docking tester; http://www.quimica.urv.cat/~pujadas/BDT), an easy-to-use graphic interface for using AutoGrid/AutoDock. BDT is a Tcl/Tk graphic front-end application that runs on top of four Fortran programs and which controls the conditions of the AutoGrid and AutoDock runs. As far as the modulation of the glucose metabolism is concerned, several in vivo and in vitro results obtained by our group have shown that grape seed procyanidin extracts (GSPE) stimulate glucose uptake in 3T3-L1 adipocytes and thus help to maintain their glucose homeostasis. In contrast, it is also well known that although some phenolic compounds do not affect glucose uptake, others reversibly inhibit it in several cell lines. Moreover, for at least some of these phenolic compounds, this inhibition is the result of their competition with ATP for the ATP-binding site in p110α (i.e. the α isoform of the catalytic subunit of phosphoinositide 3-kinase or PI3Kα). Furthermore, recent studies with isoform-specific inhibitors have identified p110α as the crucial isoform for insulin-stimulated glucose-uptake in some cell lines. Therefore, although it has been proved that the addition of phenolic compound extracts to food can have an overall benefit on health, it should be taken into account that some of these molecules may exacerbate insulin resistance in susceptible individuals via impaired glucose uptake in muscle and adipose tissues and, therefore, produce an undesirable side effect. In this context, we have applied computational approaches (i.e. protein-ligand docking and 3D-QSAR) to predict the IC50 (i.e. the concentration that reduces the p110α activity to 50%). Our results agree with previous experimental results and predict that some compounds are potential inhibitors of this enzyme. Recent results in our research group have demonstrated that the phenolic compounds in GSPE increase the activity of the farnesoid X receptor (i.e. FXR) in a dose-dependent way when the natural ligand of FXR (i.e. CDCA) is also present. The phenolic compounds might induce specific conformational changes that increase FXR activity and then contribute to cardioprotection through mechanisms that are independent of their intrinsic antioxidant capacities but that involve direct interaction with FXR to modulate gene expression. Taking into account this hypothesis a 3D-QSAR analysis was made in an attempt to understand how phenolic compounds activate FXR. So, our results explain why phenolic compounds cannot activate FXR by themselves and how they can add new interactions to stabilize the active conformation of FXR when its natural ligand (i.e. CDCA) is present. Therefore, we proposed a mechanism of FXR activation by dietary phenolic compounds in which they may enhance bile acid-bound FXR activity.

There are two chief molecular pathways to sporadic colorectal cancer (CRC), the chromosomal instability (CIN) and the CpG island methylator phenotype (CIMP) pathways. A third pathway, the pure microsatellite instability pathway, is important in inherited CRC specifically hereditary non-polyposis colorectal cancer. The CIN pathway is characterized by an adenomatous pathological precursor, aneuploidy and microsatellite stability. CIMP pathway cancers, however, are frequently proximal, develop from serrated rather than adenomatous polyps and are strongly associated with BRAF mutation. The CIMP pathway is driven primarily by epigenetic rather than genetic instability. These pathway-specific molecular traits are evident within the pathological precursors to these cancers and thus pathway divergence must occur at the beginning of carcinogenesis or even before. Although DNA methylation is recognized as a key mechanism in colorectal carcinogenesis, relatively little is known about its pattern, regulation and relevance in normal colorectal mucosa. This PhD thesis characterized the profile of DNA methylation in the normal human colorectum and explored its associations with luminal, environmental, dietary and pathological factors. The genes methylated in CRC are characterized as “type A” (Age-related) genes and “type C” (Cancer-specific) genes. Generally, “type A” genes are methylated in both normal and neoplastic tissue with the degree of methylation proportional to the age of the tissue. The methylation of “type C” genes, however, is more specific for neoplastic tissue. The primary study recruited 166 patients undergoing colonoscopy. At colonoscopy, mucosal biopsies were taken from the caecum, transverse colon, sigmoid colon and rectum. DNA methylation was analysed by MethyLight at “type A” (ESR1, GATA5, HIC1, HPP1, SFRP1) and “type C” methylation markers (MGMT, MLH1, CDKN2A, MINT2, MINT31, IGF2, CACNA1G, NEUROG1, SOCS1, RUNX3). LINE-1 methylation was quantified by pyrosequencing. The last 5 “type C” markers comprise a CIMP panel used to identify CIMP cancers. Mean “type A” and CIMP panel methylation Z-scores were calculated. The PMR for each of these CpG island loci was compared to patient age, gender, previous colorectal polyps, smoking history and the presence of concomitant pathology. Most “type A” genes demonstrated strong and direct correlations between methylation and patient age (e.g. ESR1, ρ=0.66, p<0.0001) and had greater methylation within the distal compared to the proximal colorectum (e.g. ESR1, p<0.0001). On multivariate analysis, the mucosal “type A” methylation Z-score had a strong, independent, inverse association with the diagnosis of colorectal adenomas (OR=0.23, p<0.001), the precursor to CIN cancers. The mean CIMP methylation Z-score in normal mucosa, however, was significantly and independently associated with advanced proximal serrated polyps (OR=5.1, p=0.009), the precursor to CIMP cancers. The luminal and epithelial associations with colorectal methylation were explored by a randomized, double-blind, placebo-controlled trial. This experiment was undertaken to determine whether dietary supplementation could modulate epithelial DNA methylation. In addition, the study was designed to evaluate intra-individual reproducibility of the MethyLight technique. The study consisted of a 4 week cross-over trial of resistant starch and Bifidobacterium lactis either alone or as a combined synbiotic preparation, in 20 human volunteers. Rectal biopsies, faeces and serum were collected. Rectal mucosal endpoints included DNA methylation at the CpG island loci and LINE-1, epithelial proliferation (Ki67 immunohistochemistry) and crypt cellularity. Faecal short-chain fatty acid concentrations, pH, ammonia and microbiological profiles (by DGGE and sequencing) were examined. The synbiotic intervention fostered a significantly different faecal stream bacterial community than either the prebiotic or the probiotic interventions alone, but did not show any significant associations with the epithelial or luminal parameters. To explore possible associations between luminal and epithelial parameters and mucosal DNA methylation, the baseline indices were further analysed. There was a strong positive correlation between baseline epithelial proliferation and “type A” marker methylation (ρ = 0.7, p = 0.0001). Thus, “type A” methylation may reflect the cellular age or mitotic burden of a tissue, which is a function of both time and cell turnover. There were consistent inverse trends evident between faecal short-chain fatty acid levels and rectal mucosal DNA methylation. This PhD project found that DNA methylation within the normal colorectal mucosa varied with patient age and region and was strongly associated with the development of pathway-specific pathology, suggesting that the background colorectal field may predict both the at-risk patients and at-risk pathways. Diet and the luminal environment more broadly may influence levels of DNA methylation in the colorectal mucosa and could help to explain regional patterns of colorectal DNA methylation.

Background: Tech-enabled personalised nutrition is an emerging area that has promise to improve health outcomes, widen access to nutrition expertise and reduce healthcare expenditure, yet uptake by registered dietitians remains low. This research programme aimed to identify levers and barriers that contribute to adoption of personalised nutrition in order to guide practice and policy for registered dietitians, educators and consumers. Methods: A mixed methods study with a sequential exploratory design was adopted to determine what the barriers to adoption of technologies are, and secondly, what needs to be in place to make tech-enabled personalised nutrition a reality. The research programme was conducted online using qualitative (focus groups and interviews) and quantitative measures (survey and secondary analysis). Thematic analysis, statistical and secondary analyses of data were performed respectively. Results: Using diffusion of innovation and entrepreneurial theories, findings indicate that barriers to integration of personalised nutrition technologies include intrinsic and extrinsic factors which relate to a low self-efficacy, high perception of risk, low perceived importance and usefulness of technologies to dietetic practice as well as a lack of an entrepreneurial mindset and regulatory environment. Conclusion: Uptake of tech-enabled personalised nutrition by registered dietitians will require a multi-stakeholder approach. Educational, professional, regulatory and health policies will need to be in place and strategies that open discussion between Registered Dietitians (RD’s) at all levels are needed.

Yes / Personalised nutrition (PN) has the potential to reduce disease risk and optimise health and performance. Although previous research has shown good acceptance of the concept of PN in the UK, preferences regarding the delivery of a PN service (e.g. online v. face-to-face) are not fully understood. It is anticipated that the presence of a free at point of delivery healthcare system, the National Health Service (NHS), in the UK may have an impact on end-user preferences for deliverances. To determine this, supplementary analysis of qualitative data obtained from focus group discussions on PN service delivery, collected as part of the Food4Me project in the UK and Ireland, was undertaken. Irish data provided comparative analysis of a healthcare system that is not provided free of charge at the point of delivery to the entire population. Analyses were conducted using the ‘framework approach’ described by Rabiee (Focus-group interview and data analysis. Proc Nutr Soc 63, 655-660). There was a preference for services to be led by the government and delivered face-to-face, which was perceived to increase trust and transparency, and add value. Both countries associated paying for nutritional advice with increased commitment and motivation to follow guidelines. Contrary to Ireland, however, and despite the perceived benefit of paying, UK discussants still expected PN services to be delivered free of charge by the NHS. Consideration of this unique challenge of free healthcare that is embedded in the NHS culture will be crucial when introducing PN to the UK.

No / Ultraviolet radiation (UVR) in sunlight has deleterious effects on skin, while behavioural changes have resulted in people gaining more sun exposure. The clinical impact includes a year-on-year increase in skin cancer incidence, and topical sunscreens alone provide an inadequate measure to combat overexposure to UVR. Novel methods of photoprotection are being targeted as additional measures, with growing interest in the potential for systemic photoprotection through naturally sourced nutrients. Omega-3 polyunsaturated fatty acids (n-3 PUFA) are promising candidates, showing potential to protect the skin from UVR injury through a range of mechanisms. In this review, we discuss the biological actions of n-3 PUFA in the context of skin protection from acute and chronic UVR overexposure and describe how emerging new technologies such as nutrigenomics and lipidomics assist our understanding of the contribution of such nutrients to skin health.

O consumo de frutas e verduras está relacionado com a promoção da saúde porque tem sido associado com a redução do risco de desenvolvimento de doenças crônicas como, por exemplo, câncer e doenças degenerativas e cardiovasculares. Dessa forma, o estudo dos efeitos biológicos desses alimentos tem ganhado atenção nos últimos anos. O piquiá (Caryocar villosum) é um fruto nativo da Amazônia e é rico em compostos antioxidantes como os compostos fenólicos. Assim, o objetivo deste estudo foi avaliar os efeitos genotóxicos e antigenotóxicos in vivo da polpa liofilizada do piquiá e também de seu extrato etanólico. Além disso, os compostos fitoquímicos presentes na polpa e no seu extrato foram quimicamente determinados. Ratos Wistar foram tratados por gavagem, durante 14 dias consecutivos, com três diferentes doses da polpa do piquiá (75, 150 ou 300 mg/kg p.c.) ou com seu extrato etanólico (75 mg/kg p.c.). No 14° dia, os animais receberam solução salina (NaCl 0,9%, i.p.) ou doxorrubicina (DXR, 15 mg/kg p.c., i.p.) e após 24 horas foram eutanasiados. A medula óssea e o sangue periférico foram usados no teste do micronúcleo (MN), e o fígado, rins e coração foram utilizados nos ensaios do cometa, nas análises bioquímicas das substâncias reativas ao ácido tiobarbitúrico (TBARS) e glutationa reduzida (GSH) e na avaliação da expressão de mRNA dos genes epóxido hidrolase (Ephx2) e proteína tumoral p53 (Tp53). A polpa do piquiá não apresentou efeito genotóxico nem mutagênico em nenhuma das doses avaliadas, demonstrou atividade antigenotóxica e ainda reduziu os níveis de TBARS induzidos pela DXR no coração. Efeitos opostos foram encontrados para o extrato etanólico da polpa do piquiá, por apresentar genotoxicidade, mas não mutagenicidade, e indução de TBARS no coração. Os níveis de mRNA do gene Ephx2 no rim e coração foram aumentados após o tratamento com a maior dose da polpa do piquiá, entretanto, no rim a menor dose diminuiu a transcrição desse gene induzida pela DXR. No fígado as doses de 75 e 300 mg/kg p.c. diminuíram os níveis de mRNA do gene Ephx2 induzidos pela DXR. A dose de 300 mg/kg p.c. da polpa diminuiu a expressão de mRNA do gene Tp53 nos grupos da associação piquiá + DXR no fígado, rim e coração. O extrato etanólico da polpa do piquiá modulou a expressão de mRNA do gene Ephx2 apenas no fígado, aumentando os níveis desse transcrito, enquanto que no coração houve diminuição da transcrição do gene Tp53. Foi encontrada uma diferença de composição fitoquímica entre a polpa liofilizada e seu extrato etanólico. O extrato apresentou 1,4x mais compostos fenólicos e 3x menos carotenoides quando comparado com a polpa. Além disso, o ácido gálico foi o composto fenólico predominante na polpa, enquanto que no extrato o fenol mais abundante foi o ácido elágico. A diferença dos efeitos biológicos entre a polpa liofilizada do piquiá e seu extrato etanólico pode ser devido à alteração da composição fitoquímica. / Fruit and vegetables intake has been related to the promotion of health because it has been associated to reduced risk of chronic diseases development such as cancer, and cardiovascular and degenerative diseases. Thus, the study of the biological effects of these foods has increased in recent years. Piquiá (Caryocar villosum) is a fruit native of the Amazon and it is rich in antioxidant compounds such as phenolic compounds. Therefore, the aim of this study was to evaluate the in vivo genotoxicity and antigenotoxicity effects of the piquiá lyophilized pulp fruit and its ethanolic extract. Moreover, the phytochemical characterization of pulp and extract was determined. Wistar rats were treated by gavage, for 14 days, with three doses of piquiá pulp (75, 150 or 300 mg/kg b.w.) or with its ethanolic extract (75 mg/kg b.w.). On 14th day, the animals received saline (0.9% i.p.) or doxorubicin (DXR, 15 mg/kg b.w.) and after 24 hours they were euthanized. Bone marrow and peripheral blood were used in micronucleus (MN) test, and the liver, kidney and heart were used in comet assay, thiobarbituric acid reactive substances (TBARS), reduced gluthatione (GSH), and in the evaluation of mRNA expression of epoxide hydrolase (Ephx2) and tumor protein p53 (Tp53) genes. The piquiá pulp was not genotoxic nor mutagenic, demonstrated antigenotoxic effects and reduced the TBARS levels induced by DXR in heart. The ethanolic extract had opposite effects, whereas it was genotoxic, but not mutagenic, and increased the TBARS levels in heart. Ephx2 mRNA levels in kidney and heart were increased after treatment with the higher dose of piquiá pulp, however, in kidney the lowest dose decreased the transcription of this gene induced by DXR. In liver, the 75 and 300 mg/kg b.w. doses of piquiá pulp decreased the Ephx2 mRNA levels induced by DXR. The piquiá pulp 300 mg/kg + DXR group, presented lower levels of Tp53 mRNA in liver, kidney and heart. The ethanolic extract of piquiá pulp modulated the mRNA Ephx2 expression only in the liver, increasing the levels of this transcript, while in the heart decreased the transcription of Tp53 gene. There was a difference on phytochemical composition between the pulp and its ethanolic extract. The extract presented 1.4-fold more phenolic compounds and 3-fold less carotenoids than piquiá pulp. Furthermore, gallic acid was the predominant phenol in the pulp, whereas in the ethanolic extract the most abundant phenol was the ellagic acid. The difference in the biological effects between piquiá pulp and is ethanolic extract may be due the change of the phytochemical composition.

O mercúrio é um dos metais mais nocivos presentes no ambiente, advindo tanto de fontes naturais quanto antropogênicas. Os indivíduos estão expostos a diferentes formas do mercúrio através de diversas vias, como a alimentação, principalmente no caso do metilmercúrio presente em peixes. Suas formas orgânicas são muito significativas do ponto de vista toxicológico, considerando-se a exposição da população e seus efeitos pró-oxidantes e genotóxicos envolvidos na origem de inúmeras doenças. Por outro lado, é suposto que compostos polifenólicos e outros antioxidantes da dieta podem exercer atividade protetora contra os efeitos deletérios do mercúrio. A curcumina é um pigmento amarelo polifenólico extraído do rizoma da planta Curcuma longa Linn (Zingiberaceae). Diversas evidências apontam para suas propriedades antioxidantes, de modulação da sinalização celular e alteração da expressão gênica, além da possibilidade de sua utilização na prevenção dos efeitos deletérios dos metais no organismo. Assim sendo, este trabalho teve por objetivo avaliar os efeitos da associação entre o mercúrio e a curcumina, investigando a citotoxicidade de dois compostos orgânicos de mercúrio, metilmercúrio e etilmercúrio, e da curcumina, de forma isolada e combinada, em células HepG2, utilizando o ensaio do MTT. A genotoxicidade desses mesmos compostos também foi avaliada por meio do ensaio do cometa. Além disso, a alteração no estado oxidativo das células pela razão de glutationa (GSH/GSSG) e a alteração na expressão de 84 genes relacionados com vias de dano e reparo no DNA, utilizando-se de PCR-Array, também foram avaliados. Os compostos orgânicos de mercúrio apresentaram citotoxicidade em concentrações iguais ou maiores que 16 ?M. A curcumina apresentou citotoxicidade apenas na concentração de 128 ?M. Nos experimentos de associação entre o mercúrio e a curcumina foi observado um aumento da citotoxicidade, entre a concentração de 8 ?M das duas formas de mercúrio e a concentração de 64 ?M de curcumina. Na avaliação da genotoxicidade foi observado um efeito genotóxico significativo do metilmercúrio nas concentrações de 8, 16 e 32 ?M, enquanto o etilmercúrio apresentou genotoxicidade significativa apenas na concentração de 32 ?M. A curcumina não apresentou genotoxicidade. Na associação dos compostos não foi detectada ação antigenotóxica da curcumina e houve um aumento na genotoxicidade do metilmercúrio em associação com a concentração de 32 ?M de curcumina. A razão de glutationa mostrou um aumento significativo nas células tratadas com metilmercúrio, entretanto isso não foi observado quando o metilmercúrio foi associado com a curcumina. A análise da expressão de 84 genes relacionados com danos no DNA por PCR-Array mostrou alteração na expressão de 26 genes, sendo que 3 deles tiveram aumento na expressão, DDIT3, GADD45A e PPP1R15A, relacionados principalmente com o bloqueio do ciclo celular e o processo de apoptose, e 23 genes tiveram sua expressão reduzida, relacionados com diversas vias de reparo do DNA, checkpoints do ciclo celular e apoptose. Os resultados obtidos indicam que, nas condições avaliadas, a associação da curcumina com o mercúrio aumentou os efeitos deletérios do metal, causando um aumento na citotoxicidade e na genotoxicidade do mercúrio, não se caracterizando como uma possível estratégia de prevenção dos efeitos deletérios do mercúrio / Mercury is one of the most harmful metals present in the environment arising from both natural and anthropogenic sources. The individuals are exposed to different forms of mercury through various sources, such as food, especially in the case of methylmercury in fish, with this organic forms being very significant from a toxicological point of view, considering the exposure of the population and its pro-oxidant and genotoxic effects, involved in the origin of many diseases. On the other hand it is assumed that polyphenolic compounds and other dietary antioxidants can have protective activity against the harmful effects of mercury. Curcumin is a polyphenol extracted from the rhizome of Curcuma longa Linn. (Zingiberaceae). Several evidences show its ability to act as an antioxidant, modulate cell signaling and gene expression, and the possibility of its use in chemoprevention of the deleterious effects of metals. Therefore, this study aimed to evaluate the effects of the association between mercury and curcumin, investigating the cytotoxicity of two organic compounds of mercury, methylmercury and ethylmercury, and curcumin, alone and in combination, in HepG2 cells using the the MTT assay, the genotoxicity of these same compounds through the comet assay, the changes in oxidative state of the cells by the concentration of glutathione and GSH/GSSG ratio and the changes in the expression of 84 genes related to DNA damage anda repair pathways by PCR-Array. Organic mercury compounds showed cytotoxicity at concentrations equal to or greater than 16 uM. Curcumin only showed cytotoxicity at the concentration of 128 ?M. There was an increase in cytotoxicity when mercury (8 ?M) was associated with curcumin (64 ?M). In the genotoxicity assay there was a significant genotoxic of methyl mercury at concentrations of 8, 16 and 32 ?M while ethyl mercury whas genotoxic only at 32 ?M. Curcumin was not genotoxic. There was no anti-genotoxic activity in the association of compounds and there was an increase in genotoxicity of MeHg in association with 32 ?M of curcumin. The quantification of glutathione (GSH/GSSG) showed a significant increase in the GSH/GSSG ratio in cells treated with MeHg, however this was not observed when MeHg was associated with curcumin. Gene expression analysis showed changes in the expression of 26 genes, 3 of them were upregulated, DDIT3, GADD45A and PPP1R15A, mainly related to the cell cycle blockage and apoptosis, and 23 genes were down-regulated, related with DNA repair, cell cycle checkpoints and apoptosis. These results indicate that the combination of curcumin with mercury increased the deleterious effects of the metal, causing an increase in cytotoxicity and genotoxicity of mercury, and do not represent a possible strategy to prevent the harmful effects of mercury.

Introdução: Poucos trabalhos correlacionam o perfil metabólico com o estado nutricional em crianças e adolescentes após intervenção dietética. Objetivos: Identificar, em uma população de indivíduos de 9 a 13 anos submetidos a uma intervenção nutricional, a existência de diferentes grupos metabólicos formados com base em dados bioquímicos (níveis de glicemia, colesterol total, triglicérides, VLDL colesterol, LDL colesterol e HDL colesterol) coletados em 3 momentos do estudo; e descrever a evolução longitudinal do perfil nutricional e metabólico destes grupos. Metodologia: Estudo clínico de intervenção auto-controlado, baseado na medida do perfil bioquímico (níveis de glicemia, colesterol total, triglicérides, VLDL colesterol, LDL colesterol e HDL colesterol) e do estado nutricional (antropometria, composição corporal e dados de ingestão alimentar) em três momentos: no início do estudo (antes de ser iniciada a intervenção), após seis semanas de suplementação de vitaminas e minerais e após outras seis semanas sem essa intervenção, para avaliar como um indivíduo, de 9 a 13 anos de idade responde à suplementação de múltiplos micronutrientes. O nível de atividade física praticado por esses indivíduos foi avaliado através do aparelho Bodybugg®. Resultados: Cento e trinta e seis indivíduos foram estudados até o terceiro momento do estudo. 43,4% eram do sexo masculino e 56,6% eram do sexo feminino. A média de idade foi de 11,39 ± 1,10 anos. A maioria dos participantes pertenciam ao estadiamento puberal 2 (43,4%) e 3 (35,3%). Em relação à classificação econômica dos participantes, a maioria pertencia à categoria B2 (38,2%) e C1 (26,5%). Do total da amostra estudada, no momento 1, 4,4% dos participantes apresentaram magreza grave; 5,9% apresentaram magreza, 41,9% estavam com o peso adequado, 22,8% tinham sobrepeso e 25% tinham obesidade. No momento 2, 3,7% dos participantes apresentaram magreza grave, 7,4% apresentavam magreza, 42,6% tinham o peso adequado, 22,1% tinham sobrepeso e 24,3% tinham obesidade e no momento 3, 3,7% dos participantes apresentaram magreza grave, 6,6% apresentavam magreza, 41,2% tinham o peso adequado, 22,8% tinham sobrepeso e 25,7% tinham obesidade. Em média encontramos: 3,7% dos participantes com magreza grave, 5,9% com magreza, 41,9% com peso adequado, 24,3% com sobrepeso e 24,3% com obesidade. Os participantes foram agrupados (clusterizados) utilizando-se como critério seu perfil metabólico (níveis de glicemia, colesterol total, triglicérides, VLDL colesterol, LDL colesterol e HDL colesterol) nos três momentos do estudo, por meio da técnica estatística K-cluster. O cluster 1 (n = 111) era composto por mais indivíduos do sexo feminino que o cluster 2 (n = 25) (p = 0,006) e apresentou melhor perfil metabólico (melhores valores para o perfil lipídico e de glicemia). Os indivíduos do cluster 1 também apresentaram menor peso, índice de massa corporal (IMC), circunferência da cintura (CC) e massa gorda (% peso) quando comparado aos participantes do cluster 2. A massa corporal magra (% peso) e a água corporal total (% peso) foram estatisticamente maiores nos participantes do cluster 1. A análise da ingestão habitual (por questionário de frequência alimentar - QFA) mostrou que os participantes do cluster 1 estavam ingerindo mais vitamina B2 e vitamina B6 quando comparados aos participantes do cluster 2 (p < 0.05). Houve menores valores para proteína C-reativa (PCR) e maiores valores para ferro sérico no cluster 1. A capacidade latente de ligação de ferro (UIBC) e leucócitos foram estatisticamente maiores no cluster 2. Não houve diferença entre o nível de atividade física praticado pelos dois clusters, ambos desempenhavam atividade física leve. A análise longitudinal mostrou que houve aumento de estatura e peso nos clusters 1 e 2. A avaliação longitudinal da ingestão habitual (QFA) no cluster 1 mostra redução da ingestão de energia, carboidrato, proteína e lipídio do momento 1 (M1) para momento 2 (M2) e momento 3 (M3). A suplementação de vitaminas mostrou resultados estatísticos significativos, consistentes com suplementação e wash out para a maioria das vitaminas e minerais nos clusters 1 e 2. A análise longitudinal (corrigindo para as variáveis idade, gênero, estadiamento puberal e ingestão de energia, carboidrato e lipídio) no cluster 1 mostrou que o colesterol total e a LDL diminuíram ao longo do estudo; a glicemia diminuiu do momento 1 para o momento 2, porém PCR aumentou no momento 2; ferro sérico e hemoglobina diminuíram no momento 2 e aumentaram no momento 3. No cluster 2, o colesterol total e o LDL diminuíram ao longo do estudo; a PCR aumentou ao longo do estudo, ferro sérico diminuiu do momento 1 para o momento 2. Conclusões: Foram encontrados dois grupos metabólicos opostos. Os indivíduos podem responder de forma diferente a uma mesma intervenção e é possível que a suplementação de múltiplos micronutrientes tenha um papel na melhora do perfil glicídico e lipídico de alguns sujeitos do estudo. Estudos de genotipagem e proteômica poderão reforçar esta hipótese e ajudar a entender como o sistema biológico de crianças e adolescentes interage para culminar em uma resposta frente a uma intervenção. / Introduction: Few studies have correlated metabolic profile and nutritional status in children and adolescents after dietary intervention . Objectives: To identify the existence of different metabolic groups formed by individuals 9-13 years undergoing nutritional intervention through biochemical data (glucose levels, total cholesterol, triglycerides, VLDL cholesterol, LDL cholesterol and HDL cholesterol) collected in three stages of the study, and to describe the longitudinal evolution of the nutritional and metabolic profile in these groups. Methodology: Clinical intervention self-controlled study, based on measuring the biochemical profile (glucose levels, total cholesterol, triglycerides, VLDL cholesterol, LDL cholesterol and HDL cholesterol) and nutritional status (anthropometry, body composition and dietary intake data) at three times: at baseline (before the intervention started), after six weeks of supplementation of vitamins and minerals and after more six weeks without this intervention, to assess how an individual, 9-13 years old answered the multiple micronutrient supplementation. Physical activity level was also assessed through a tool called bodybugg®. Results: One hundred and thirty six subjects were studied until the third moment of the data collection. 43.4% were male and 56.6% were female. The average age was 11.39 ± 1.10 years. Most participants belonged to pubertal stage 2 (43.4%) and 3 (35.3%). Regarding the economic status of the participants, the majority belonged to the category B2 (38.2%) and C1 (26.5%). Of the total sample, at moment 1, 4.4% of participants had severe underweight, 5.9% were underweight, 41.9% were with the proper weight, 22.8% were overweight and 25% were obese. At the moment 2, 3.7% of participants had severe underweight, 7.4% were underweight, 42.6% had normal weight, 22.1% were overweight and 24.3% were obese. And at the moment 3, 3.7% of participants had severe thinness, 6.6% were underweight, 41.2% had normal weight, 22.8% were overweight and 25.7% were obese. On average we found: 3.7% of participants with severe thinness, 5.9% with malnutrition, 41.9% with adequate weight, 24.3% overweight and 24.3% obese. The clustering of the participants used as criteria the metabolic profile (glucose levels, total cholesterol, triglycerides, VLDL cholesterol, LDL cholesterol and HDL cholesterol) of the three stages of the study through the statistical approach K - cluster. Cluster 1 (n = 111) had a higher proportion of females when compared to cluster 2 (n = 25) (p = 0.006) and better metabolic profile (lipid and glycemia). Participants who had better metabolic profile (cluster 1) showed lower weight, body mass index (BMI), waist circumference (WC) and fat mass (weight %) when compared to participants in cluster 2. Lean body mass (weight %) and total body water (% weight) were statistically higher in participants in cluster 1. The analysis of the habitual intake (food frequency questionnaire - FFQ) showed that participants in cluster 1 were had higher intake of vitamin B2 and vitamin B6 when compared to cluster 2 (p < 0,05). Cluster 1 also showed higher values C - reactive protein (CRP) and higher levels of iron. The latent capacity for iron binding (UIBC) and leukocytes were higher in cluster 2. There was no difference between the level of physical activity practiced by clusters 1 and 2, both had light physical activity. Longitudinal analysis showed that there was an increase in height and weight in clusters 1 and 2. A longitudinal assessment of usual intake (FFQ) in cluster 1 showed reduced intake of energy, carbohydrate, protein and lipid from moment 1 (M1) to moment 2 (M2) and moment 3 (M3). Supplementation with vitamins showed significant statistical results, consistent with supplementation and washout for most vitamins and minerals in clusters 1 and 2. A longitudinal analysis (correcting for age, gender, pubertal stage, and energy intake, carbohydrate and lipid) in cluster 1 showed that total cholesterol and LDL decreased throughout the study, blood glucose decreased from time 1 to time 2, but CRP increased in moment 2; hemoglobin and serum iron decreased from 1 to 2 and from 1 to 3. In cluster 2, total cholesterol and LDL decreased throughout the study, CRP increased throughout the study, serum iron decreased from time 1 to time 2. Conclusions: We found two reverse metabolic groups. Individuals may respond differently to the same intervention, and it is possible that multiple micronutrient supplementation has a role in improving glycemia and lipid profile of some subjects. Genotyping and proteomics studies may reinforce this hypothesis and help understand how the biological system of children and adolescents interact to culminate in a response against an intervention.